In the yeast Saccharomyces cerevisiae, the most important systems for conveying excess cytosolic NADH to the mitochondrial respiratory chain are the external NADH dehydrogenases (Nde1p and Nde2p) and the glycerol-3-phosphate dehydrogenase shuttle. In the latter system, NADH is oxidized to NAD+ and dihydroxyacetone phosphate is reduced to glycerol-3-phosphate by the cytosolic Gpd1p. Subsequently, glycerol-3-phosphate donates electrons to the respiratory chain via mitochondrial glycerol-3-phosphate dehydrogenase (Gut2p). At saturating concentrations of NADH, the activation of external NADH dehydrogenases completely inhibits glycerol-3-phosphate oxidation. Studies on the functionally isolated enzymes demonstrated that neither Nde1p nor Nde2p directly inhibit Gut2p. Thus, the inhibition of glycerol-3-phosphate oxidation may be due to competition for entrance of electrons into the respiratory chain. Using single deletion mutants of Nde1p or Nde2p, we have shown that glycerol-3-phosphate oxidation via Gut2p is fully inhibited when NADH is oxidized via Nde1p, whereas only 50% of glycerol-3-phosphate oxidation is inhibited when Nde2p is functioning. By comparing respiratory rates with different respiratory substrates, we show that : (i) electrons from Nde1p are favored over electrons coming from Ndip (internal NADH dehydrogenase); (ii) when electrons are coming from either Nde1p or Nde2p, and succino-dehydrogenase, their use by the respiratory chain is shared up to a comparable extent. This suggests a very specific competition for electron entrance into the respiratory chain, which may be due to supramolecular organization of the respiratory chain. The physiological consequences of such regulation are discussed.